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Abstract

Bacterial soft rot is a destructive disease affecting various ornamental plants and crops worldwide. Causative bacteria are frequently identified as members of the genus Dickeya. Soft rot disease limits orchid production and causes severe economic losses in orchid nurseries globally. At the moment no effective chemical-based approach is available to control or treat Dickeya infections. Therefore, the bacteriophage biocontrol approach represents a promising management strategy for preventing and controlling Dickeya soft rots. Within this doctoral thesis, we aimed to comprehensively characterise and taxonomically classify causative pathogenic bacteria from symptomatic Phalaenopsis orchids from a commercial production site, as well as to isolate and characterise bacteriophages against bacterial isolates and assess their biocontrol potential.
In previous experiments, the most commonly isolated bacteria from diseased Phalaenopsis tissue were identified as Dickeya spp. Genetic diversity divided the isolates into two groups, with the greatest similarity to previously reported undefined Dickeya lineages from orchids (UDL-3 and UDL-4). Two model isolates, Dickeya sp. B16 (UDL-3) and S1 (UDL-4), representing the observed diversity, were comprehensively characterised. Virulence and aggressiveness comparison tests in vivo on Phalaenopsis orchids, potato plants and witloof chicory leaves indicated the high virulence and extreme maceration potential of these novel Dickeya isolates, compared to a reference panel of Dickeya spp.
In the last decade, the genus Dickeya has undergone multiple changes as multiple new taxa have been described, and just recently, a new putative Dickeya species was reported, namely, Dickeya fangzhongdai. However, Dickeya sp. B16 and S1 genetically and genomically corresponded to a, at the time, unclassified group of Dickeya strains isolated from monocotyledonous plants and water. Average nucleotide identity analysis and multilocus sequence analysis enabled us to classify these isolates as the new D. fangzhongdai species. However, variations among the isolates and D. fangzhongdai type strain could be observed on phenotypic level.
Bacteriophages active against causative Dickeya isolates were isolated from sewage water demonstrating the applicability of the convective interaction media monolith chromatography for bacteriophage isolation from environmental sources. Bacteriophages isolated from sewagewater and previously isolated bacteriophages from rotted Phalaenopsis tissue were classified into 3 different families, Podoviridae, Myoviridae and Siphoviridae, expressing slight variations in their host profiles. Bacteriophage BF25/12 from the Podoviridae family was comprehensively characterised (pH, UV and temperature stability, resistance development and whole-genome analysis), and molecular tools for tracking and quantification were developed and initial bacteriophage–host interactions were examined. Therefore, we have successfully established a bacteriophage–host system for studying its dynamics that is essential for effective bacteriophage applicability.
The results of the research work presented in this thesis represent the first comprehensive characterisation of Dickeya isolates corresponding to the D. fangzhongdai species. Furthermore, we have established a well-defined system for a bacteriophage-host dynamics study, a first for the genus Dickeya.